The enzymically active order complex resembling horse heart cytochrome c can be reconstituted from heme fragment (1-25)H (residues 1 to 25) and apofragments (28-38) and (56-104) within 1 s with apparent dissociation constant less than 10 to minus the 5th power and less than 10 to minus 15th power M2 in the ferric and the ferrous form, respectively, at pH 7.0 and 25 degrees C. Our observations have suggested a pathway in assembly of the ferric complex: (1-25)H (56-104) (1225)H:(56-104) (28-38) (1-25)H:(28-38):(56-104). Ligation of Met 80 to the heme iron seems to occur in a late phase of folding of this complex. In order to investigate the stabilizing force of this complex using the fragment exchange technique, Met-65 and 80 of fragment (56-104) was labeled, using (H3)CH3I. While the dissociation rate of non-productive complex ferrous (1-25)H:(56-104) is very fast (half-life, less than 20 s at 0-20 degrees C), the dissociation rate of labeled (56-104) from the three fragment complex ferrous (1-25)H:(28-38):(56-104) is slow and highly dependent on temperature (1.5 x 10 to minus 5th power, 5 x 10 to minus 5th power, 1 x 10 to minus 4th power, 4.2 x 10 to minus 4th power and 5.6 x 10 to minus 3rd power s-1 at 15 degrres, 20 degrees, 25 degrees, 30 degrees and 37 degrees C, respectively) indicating that cooperative interatomic interactions underlie the stabilizing force.